The role of cytomegalovirus antiapoptotic proteins in replication in macrophages.

描述（由申请人提供）：人巨细胞病毒（HCMV）是一种高度流行的疱疹病毒，可通过输血或移植有效传播。通过白细胞减少和病原体灭活技术的灭菌技术（例如，pk10/UVA），但开发用于移植的HCMV安全细胞疗法需要更复杂的策略。作为设计依赖于减少病毒控制细胞死亡的策略的长期目标的一部分，该提案探讨了病毒细胞死亡抑制基因UL 36在巨噬细胞中HCMV生长中的作用，目的是验证UL 36作为细胞治疗组分中选择性抑制HCMV的靶标。单核细胞和源自它们的巨噬细胞非常可能是基于以下的这种操作的靶细胞：1）粒细胞、巨噬细胞和树突状细胞（GM-Ps）的祖细胞是HCMV传播的重要贡献者，因为它们携带作为感染源的潜伏病毒，2）这些祖细胞分化为巨噬细胞允许病毒的溶解性生长，和3）巨噬细胞中的生长被认为是HCMV传播和随后疾病的关键步骤。鼠CMV（MCMV）UL 36同源物M36对于巨噬细胞中的复制是必需的，并且预期HCMV中的UL 36对于巨噬细胞中的生长也是必需的。预计UL 36的结合半胱氨酸天冬氨酸蛋白酶原-8和防止活化的性质也使其在产生容许细胞的分化过程中成为有吸引力的靶标，因为这种蛋白酶在经历巨噬细胞分化的单核细胞中以及在凋亡期间是重要的。此外，已经发现了由UL 36基因编码的新的细胞死亡抑制功能，其扩展了该基因在这种重要细胞类型中的预期作用。将启动长期目标，并在两个特定目标中解决近期目标，这两个特定目标旨在1）确定差异剪接对巨噬细胞中UL 36基因功能的贡献，以及2）确定microRNA miR 36-1对巨噬细胞中UL 36基因功能的贡献。重组病毒和强大的，基于转染的策略将被纳入一个集中的努力，以确定UL 36基因产物，抑制多个细胞死亡途径部署的巨噬细胞抑制HCMV复制。公共卫生相关性：人巨细胞病毒（CMV）是一种高度流行的疱疹病毒，可导致危及生命的疾病，特别是在输血、移植后，或者如果它传播给未出生的孩子。CMV安全的细胞移植疗法的发展需要新的方法;这些策略的发展可能会表明在其他情况下预防传播的有效手段。这项提案将确定病毒机制，防止内在防御杀死感染细胞重要的疾病传播。

DESCRIPTION (provided by applicant): Human cytomegalovirus (HCMV) is a highly prevalent herpesvirus efficiently transmitted by transfusion or transplantation. Sterilization techniques through leukoreduction and pathogen inactivation techniques (e.g., psoralen/UVA) are available but the development of HCMV-safe cellular therapies for transplantation requires more sophisticated strategies. As part of a long-term goal to devise strategies that rely on reducing viral control of cell death, this proposal explores the role of the viral cell death suppression gene, UL36, in HCMV growth in macrophages, with the objective of validating UL36 as a target for selectively suppressing HCMV in cellular therapy components. Monocytes and the macrophages derived from them are highly likely target cells for this manipulation based on the following: 1) progenitors of granulocytes, macrophages, and dendritic cells (GM-Ps) are important contributors to HCMV transmission because they harbor latent virus that serves as the source for infection, 2) differentiation of these progenitors to macrophages allows for lytic growth of the virus, and 3) growth in macrophages is considered to be a critical step for HCMV transmission and ensuing disease. The murine CMV (MCMV) UL36 homolog, M36, is essential for replication in macrophages, and it is anticipated that UL36 in HCMV will also be essential for growth in macrophages. It is anticipated that properties of UL36, binding procaspase-8 and preventing activation, also makes it an attractive target during the differentiation process that yields permissive cells because this protease is important in monocytes undergoing macrophage differentiation as well as during apoptosis. Further, novel cell death suppression functions encoded by the UL36 gene have been uncovered that expand the anticipated role of this gene in this important cell type. Long-term goals will be initiated and immediate goals addressed in two specific aims designed to 1) determine the contribution of differential splicing to UL36 gene function in macrophages, and 2) determine the contribution of the microRNA miR 36-1 to UL36 gene function in macrophages. Recombinant viruses and robust, transfection-based strategies will be incorporated in a focused effort to identify the UL36 gene products that inhibit multiple cell death pathways deployed by macrophages to inhibit HCMV replication. PUBLIC HEALTH RELEVANCE: Human cytomegalovirus (CMV) is a highly prevalent herpes virus that causes life- threatening disease, especially following blood transfusions, transplantations, or if it is transmitted to an unborn child. The development of CMV-safe cellular therapies for transplantation requires novel approaches; the development of these strategies will likely indicate effective means of preventing transmission in other situations. This proposal will determine viral mechanisms that prevent intrinsic defenses that kill infected cells important to disease transmission.